The purpose of this grant is to provide the Principle Investigator with stable longterm salary support to allow her to further develop her research career. The availability of such support would permit the applicant to substantially reduce her administrative and service load and to concentrate on the experimental program described herein. Short-term goal (during the period of support) are to clarify the role of calcium binding at contraction in striated muscle. Long-term goals include enhancement of the candidate's standing as an independent investigator in the areas of striated muscle regulation and ultrastructure (with particular emphasis on cardiac muscle) and computer-aided image analysis. The Center for Bioengineering provides a strong research and educational environment for the attainment of these goals and the state-of-the-art equipment required for the project are in place and fully functional. Collaborations within and outside of the Center provide an excellent opportunity for further training and fruitful research. The proposed research will apply analytical and structural techniques to mammalian skeletal and cardiac muscle in order to study several aspects of the role of calcium in activation and contraction. Electron probe microanalysis will be used to measure calcium sequestered at junctional sites of the sarcoplasmic reticulum in rat and ferret papillary muscles under a variety of conditions, with the long-term objective being to understand the relative role of intra- and extracellular calcium in cardiac contractility. Related experiments on skeletal and cardiac skinned fibers will address the role of calcium bound to myofilaments. The electron probe will be used to measure crossbridge state and sarcomere length in A, I and overlap regions. The use of digital structural analysis of skinned and intact muscle samples using freeze fracture deep etch replicas will address the question of whether structural changes in the myosin filaments are associated with calcium binding and activation. Since many endogenous and clinically used agents are thought to modify cardiac output via changes in calcium, these results are potentially very important in understanding the normal and pathological functioning of the heart.